Effects of Child and Maternal Histo-Blood Group Antigen Status on Symptomatic and Asymptomatic Enteric Infections in Early Childhood

Funding Information: Financial support. This work was funded by the Etiology, Risk Factors, and Interactions of Enteric Infections and Malnutrition and the Consequences for Child Health and Development Project (MAL-ED) is carried out as a collaborative project funded by the Bill & Melinda Gates Foundation (BMGF) (BMGF-47075), the Foundation for the National Institutes of Health, and the National Institutes of Health, Fogarty International Center, whereas additional support was obtained from BMGF for the examination of host innate factors on enteric disease risk and enteropathy (Grants OPP1066146 and OPP1152146; to M. N. K.). Additional funding was obtained from teh Sherrilyn and Ken Fisher Center for Environmental Infectious Diseases, Johns Hopkins School of Medicine (to M. N. K) and the National Center for Research Resources and the National Center for Advancing Translational Sciences, National Institues of health 1UL1TR001079. Acknowledgments. We thank the participants, their families, and the study community for their dedicated time and effort to better the understanding the transmission and more enduring impact of enteric infections in early childhood. We also thank the following: Jan Vinje (Centers for Disease Control and Prevention) for critical input and manuscript review; Dr. Leah Jager for consultation regarding the statistical analysis; Dr. Ben Jann (University of Bern, Switzerland) for guidance in generating the figures; Christine Szymanski for insight and encouragement, particularly regarding Campylobacter infection and disease patency; Chris Damman and Anita Zaidi for input on early iterations of the analysis; and Dick Guerrant for final reflections.Peer reviewe

The impact of antigen processing on CD8+ t cell memory inflation

T cell "memory inflation" is the sustained induction of effector memory cells that home to peripheral tissues and retain their functionality. Defining the mechanisms that drive these non-classical memory responses may contribute towards the development of novel prophylactic or therapeutic vaccines. A model of memory inflation based on responses to β-galactosidase delivered by a non-replicating adenoviral vector provides a robust tool for investigating the underlying mechanisms. This work has shown that these responses are not dependent upon the human cytomegalovirus (HCMV) promoter within the model, this being the only part of the model that is CMV-derived. This model has been used to test the hypothesis that bypassing antigen processing would result in inflationary memory responses to CD8+ T cell epitopes that are not normally the targets of such responses. When the β-gal497-504 restricted epitope (ICPMYARV) was expressed as a minigene in a recombinant adenovirus vector, inflationary CD8+ T cell responses were induced, instead of the classical responses obtained with full-length β-galactosidase. Similar results were obtained with the M45985-993 (HGIRNASFI) epitope from the mouse cytomegalovirus M45 protein. These data demonstrate that the polypeptide context of a CD8+ T cell epitope may determine whether classical or inflating memory responses are induced. This could be relevant to the design of recombinant antigens in adenoviral vectors, which have emerging therapeutic and prophylactic applications.</p

Complete Genome Sequence of Vibrio campbellii DS40M4

We present the complete genome sequence of Vibrio campbellii DS40M4, assembled from Illumina and Oxford Nanopore data. This effort improves upon a previous draft assembly to resolve this organism’s two-chromosome and one-plasmid genetic structure and to provide valuable context for evaluating the gene arrangement and evolution of this species

Adenoviral vaccines promote protective tissue-resident memory T cell populations against cancer

Background Adenoviral vectors emerged as important platforms for cancer immunotherapy. Vaccination with adenoviral vectors is promising in this respect, however, their specific mechanisms of action are not fully understood. Here, we assessed the development and maintenance of vaccine-induced tumor-specific CD8+ T cells elicited upon immunization with adenoviral vectors.Methods Adenoviral vaccine vectors encoding the full-length E7 protein from human papilloma virus (HPV) or the immunodominant epitope from E7 were generated, and mice were immunized intravenously with different quantities (107, 108 or 109 infectious units). The magnitude, kinetics and tumor protection capacity of the induced vaccine-specific T cell responses were evaluated.Results The adenoviral vaccines elicited inflationary E7-specific memory CD8+ T cell responses in a dose-dependent manner. The magnitude of these vaccine-specific CD8+ T cells in the circulation related to the development of E7-specific CD8+ tissue-resident memory T (TRM) cells, which were maintained for months in multiple tissues after vaccination. The vaccine-specific CD8+ T cell responses conferred long-term protection against HPV-induced carcinomas in the skin and liver, and this protection required the induction and accumulation of CD8+ TRM cells. Moreover, the formation of CD8+ TRM cells could be enhanced by temporal targeting CD80/CD86 costimulatory interactions via CTLA-4 blockade early after immunization.Conclusions Together, these data show that adenoviral vector-induced CD8+ T cell inflation promotes protective TRM cell populations, and this can be enhanced by targeting CTLA-4

Effect of acute MCMV infection on pre-existing CD8⁺ D8V⁺ memory cells.

<p>(A) Mice were first immunized with Ad-lacZ, then infected >50 days later with MCMV. Data are combined from two independent experiments (N = 7–9 mice per group). Peripheral lymphocytes were sampled at 21 days after MCMV infection and stained <i>ex vivo</i> with CD8, CD44 and CD62L. The proportions of effector memory (CD44⁺ CD62L low), central memory (CD44⁺CD62L⁺) and naïve (CD44low CD62L⁺) was determined. (B) Levels of D8V-specific tetramer population was measured between days 2–7 after infection with MCMV in the blood (N = 4–5 from two independent experiments) and at day 4 in the spleen, bone marrow, lymph nodes and liver. (C) <i>In vivo</i> CTL assay. Splenocytes from mice immunized with Ad-lacZ 45 days previously were isolated and labelled with CFSE then equal numbers of CFSE-labelled splenocytes were adoptively transferred into mice infected with MCMV at day 1 post-infection or a group of uninfected controls. The levels of transferred D8V⁺ effecter memory cells was followed in the blood over time by <i>ex vivo</i> tetramer staining (N = 6 per group from two independent experiments). To ensure equal numbers of splenocytes were transferred, the percentage of CFSE⁺CD8⁺ cells in naïve and MCMV-infected recipients were compared at the indicated time points. (D) The levels of the inflating effector memory D8V tetramer⁺ population was measured in naïve mice and a group acutely infected with MCMV at the indicated time points. p values were measured by Mann-Whitney test. *p<0.05.</p

Sequential infection with MCMV followed by Ad-LacZ.

<p>(A) C57BL/6 mice were first infected with MCMV, then >50 days later were infected with Ad-lacZ. The size of the pre-existing MCMV specific M38 effector memory response after Ad-lacZ infection was measured in the blood and liver by <i>ex-vivo</i> tetramer staining. The percentage of pre-existing M38 response in the blood and (B) the absolute numbers in the liver after immunization with Ad-lacZ vector was determined at the indicated time points. (C) The kinetics and magnitude of the new inflating response (D8V) in groups of mice with latent MCMV was also measured by <i>ex vivo</i> tetramer staining compared against uninfected mice. The percentage of D8V tetramer+ cells in the blood and (D) the absolute numbers in the liver was measured at the indicated time points. The figures show the mean from 3–8 mice per time point obtained from 2 independent experiments. p values were measured by Mann-Whitney tests. *p<0.05.</p

Sequential infection with Ad-LacZ followed by MCMV.

<p>C57BL/6 mice were first immunized with Ad-lacZ, then >50 days later were infected with MCMV. The size of the pre-existing Ad-lacZ specific M38 effector memory response after MCMV infection was measured in the blood and liver by ex-vivo tetramer staining. (A) The percentage of pre-existing inflating D8V tetramer+ population in the blood and (B) the absolute numbers in the liver after MCMV infection was determined at the indicated time points. (C) The kinetics and magnitude of the new inflating response (M38) in groups of mice previously immunized with Ad-lacZ was also measured by <i>ex vivo</i> tetramer staining and compared against uninfected mice. The percentage of M38 tetramer⁺ cells in the blood and (D) the absolute numbers in the liver was measured at the indicated time points. The figures show the mean from 3–8 mice per time point obtained from 2 independent experiments. p values were measured by Mann-Whitney tests followed by Dunn’s multiple comparison. *p<0.05, **p<0.005.</p

Adenovirus vector vaccination reprograms pulmonary fibroblastic niches to support protective inflating memory CD8+ T cells.

Pathogens and vaccines that produce persisting antigens can generate expanded pools of effector memory CD8+ T cells, described as memory inflation. While properties of inflating memory CD8+ T cells have been characterized, the specific cell types and tissue factors responsible for their maintenance remain elusive. Here, we show that clinically applied adenovirus vectors preferentially target fibroblastic stromal cells in cultured human tissues. Moreover, we used cell-type-specific antigen targeting to define critical cells and molecules that sustain long-term antigen presentation and T cell activity after adenovirus vector immunization in mice. While antigen targeting to myeloid cells was insufficient to activate antigen-specific CD8+ T cells, genetic activation of antigen expression in Ccl19-cre-expressing fibroblastic stromal cells induced inflating CD8+ T cells. Local ablation of vector-targeted cells revealed that lung fibroblasts support the protective function and metabolic fitness of inflating memory CD8+ T cells in an interleukin (IL)-33-dependent manner. Collectively, these data define a critical fibroblastic niche that underpins robust protective immunity operating in a clinically important vaccine platform

Effect of boosting with Ad-lacZ on the depleted responses.

<p>(A) C57BL/6 mice were first immunized with Ad-lacZ and then >50 days later were infected with MCMV i.v. After >50 days post- MCMV infection, the mice were boosted with a second dose of Ad-lacZ i.v. The levels of CD8⁺D8V⁺ in the blood was measured by <i>ex vivo</i> tetramer staining after primary Ad-lacZ infection. The data shown are from one of two independent experiments (N = 3). (B) The distribution of the boosted cells in non-lymphoid organs were measured at day 50 after 2^nd dose of Ad-LacZ. (C) The proportion of CD44⁺ CD62L^- expression in <i>ex vivo</i> peripheral blood 6 days after primary Ad-lacZ immunization, and 6 days after second dose of Ad-lacZ. (N = 4–6 mice from 2 independent experiments). (D) The figures show the proportion of CD44⁺CD62L^- (left column) and CD27^-CD62L^- (right column) expression in CD8⁺D8V⁺ cells from the blood (upper row) and liver (lower row) of Ad-lacZ only, Ad-lacZ+MCMV and Ad-lacZ+MCMV boosted with Ad-lacZ groups, as measured by ex-vivo staining. The data are from two or more independent experiments (N = 4–11). p values were measured by two-way Anova followed by Sidak’s multiple comparison test. *p<0.05, **p<0.005.</p